Myocardial infarction (irreversible damage to heart tissue, often due to heart attack) is a common life-threatening event that may cause sudden death or heart failure. The ventricular dysfunction that arises after myocardial infarction results, primarily, from a massive loss of cardiomyocytes and gradual replacement of damaged cardiomyocytes with fibrotic non-contractile (scar) tissue. In most cases, the loss of cardiomyocytes after myocardial infarction is irreversible. Indeed, it is widely accepted that the proliferative (and, therefore, the regenerative) potential of adult mammalian cardiomyocytes is quite limited (Rumyantsev and Carlson, Growth and Hyperplasia of Cardiac Muscle Cells (New York: Harwood Academic Publishers, 1991)), although this view has recently been challenged (Leri et al., Mol. Cell. Cardiol., 3:385-90, 2000; Kajstura et al., Am. J. Pathol., 156:813-19, 2000; Beltrami et al, N. Engl. J Med., 344(23):1750-57, 2001).
Despite considerable advances in the diagnosis and treatment of heart disease, cardiac damage and dysfunction relating to myocardial infarction are still among the major cardiovascular disorders. Accordingly, it remains a major therapeutic challenge to find new effective approaches to improve cardiac function after myocardial infarction.
The potential to reactivate cardiomyocyte proliferation through the manipulation of putative cellular regulators, or the conversion of pluripotent stem cells to cardiomyocytes (Orlic et al., Nature, 410:701-05, 2001), offers an exciting impetus for the design of novel therapeutic interventions to enhance cardiac function during disease conditions. The bulk of evidence obtained over the past decade maintains, however, that mammalian cardiomyocytes proliferate throughout fetal development and into the early neonatal period, at which time DNA replication declines quickly and cell division ceases (Beinlich and Morgan, Mol. Cell. Biochem., 119:3-9, 1993; Casscells et al., J. Clin. Invest., 85:433-41, 1990; Speir et al., Circ. Res., 71:251-59, 1992; Parker and Schneider, Annu. Rev. Physiol., 53:179-200, 1991; Simpson, P. C., Annu. Rev. Physiol., 51:189-202, 1989). Transition from hyperplastic growth (cell division) to hypertrophic growth (increase in cell size) then ensues. In the murine heart, cardiomyocyte division is reportedly completed by birth, with DNA synthesis in neonatal cells (through post-natal day 3) contributing only to binucleation (Soonpaa et al., J. Mol. Cell. Cardiol., 28:1737-46, 1996). The cessation of myocyte proliferation is attributed to an arrest of the cell cycle (Brooks et al., Cardiovasc. Res., 39:301-11, 1998). In accordance with this hypothesis, adult rat cardiomyocytes have been shown to display a dual cell-cycle blockade, with approximately 80% of cells arresting in G0/G1, and 15% -20% of cells arresting in G2/M (Poolman and Brooks, Mol. Cell. Cardiol., 29:A19 (Abstract), 1997; Poolman et al., Int. J. Cardiol., 67:133-42, 1998).
Progression through the cell cycle is tightly regulated, and involves cyclins complexed with their catalytic partners, the cyclin-dependent kinases (cdks). Among the cyclins, cyclin A2 is unique in that it regulates progression through two critical transitions: cyclin A2 complexed with cdk2 is essential for the G1/S transition, and cyclin A2 complexed with cdk1 promotes entry into mitosis (Sherr and Roberts, Genes Dev., 9:1149-63, 1995; Pagano et al., EMBO J., 11:961-71, 1992). It is well-established that mammalian cardiomyocytes cease to proliferate in the early neonatal period due to arrest of the cell cycle. Cyclin A2 is the only cyclin to be completely downregulated, at both the message and protein level, during cardiogenesis, in rat and human, in a manner that appears coincident with this withdrawal of cardiomyocytes from the cell cycle (Yoshizumi et al., J. Clin. Invest., 95:2275-80, 1995).
Previously, it has been shown that zebrafish fully regenerate hearts within 2 months of 20% ventricular resection, due to robust proliferation of cardiomyocytes localized at the leading epicardial edge of the new myocardium. This injury-induced cardiomyocyte proliferation was able to overcome scar formation, allowing cardiac muscle regeneration. It has been suggested that this regeneration of heart tissue in zebrafish is related to the Mps1 mitotic checkpoint kinase (Poss et al., Heart regeneration in zebrafish. Science, 298:2188-90, 2002). It has also been shown that cardiomyocytes react to myocardial infarction by activating cyclins and cyclin-dependent kinases (Reiss et al., Myocardial infarction is coupled with activation of cyclins and cyclin-dependent kinases in myocytes, Exp. Cell Res., 225:44-54, 1996). However, prior to the present invention, it had not been directly demonstrated that regulation of cyclins, particularly cyclin A2, can induce cardiomyocyte mitosis once the timeline for cell-cycle exit (and, therefore, “terminal” differentiation) has been surpassed.